ABSTRACT Nonsyndromic orofacial clefts (OFCs) are genetically complex structural birth defects caused by genetic factors, environmental exposures, and their interactions. OFCs are the most common craniofacial anomalies in humans, affecting approximately 1 in 700 newborns, and are one of the most common structural birth defects worldwide. On average a child with an OFC initially faces feeding difficulties, undergoes 6 surgeries, spends 30 days in hospital, receives 5 years of orthodontic treatment, and participates in ongoing speech therapy, leading to an estimated total lifetime treatment cost of about $200,000. Further, individuals born with an OFC have higher infant mortality, higher mortality rates at all other stages of life, increased incidence of mental health problems, and higher risk for other disorders (notably including breast, brain, and colon cancers). Genome-wide linkage and association studies by our group and others have identified approximately 18 genomic regions likely to contribute to the risk for nonsyndromic OFCs which together account for about 55-60% of the heritability for OFC. Despite this substantial progress, the functional/pathogenic variants at OFC-associated regions are mostly still unknown. Because previous OFC genomic studies (genome-wide linkage, genome-wide association studies (GWAS), and targeted sequencing) are based on relatively sparse genotyping data, they cannot distinguish between causal variants and variants in linkage disequilibrium with unobserved causal variants. Moreover, it is unknown whether the association or linkage signals are due to single common variants, haplotypes of multiple common variants, clusters of multiple rare variants, or some combination. Part of the ?missing heritability? for OFC may be accounted for by rare variants within previously associated genomic regions. Finally, we cannot yet attribute specific genetic risk to individual cases and case families. Therefore, the primary goal of the current study is identify specific OFC risk variants by analyzing whole genome sequencing (WGS) of 415 OFC parent-case trios. The WGS of the trios is provided from our Kids First project (X01-HL132363 ?Kids First: Genomic Studies of Orofacial Cleft Birth Defects?), part of the Gabriella Miller Kids First Pediatric Research Program, a trans- NIH effort currently focused on gene discovery (https://commonfund.nih.gov/KidsFirst),in pediatric cancers and structural birth defects. Our analyses have the following specific aims: (i) to comprehensively interrogate the genetic architecture of known OFC- associated regions; (ii) to identify new variant associations with OFC.